|
|
|
| Isolation and Degradation Characteristics of Cellulose-degradation Bacteria at Room and Low Temperature |
| MENG Jian-Yu, CHEN Wu-Li-Ji-Ma, GUO Hui-Qin, FENG Fu-Ying, CHEN Yu-Ping* |
| College of Life Sciences, Inner Mongolia Agricultural University, Hohhot 010011, China |
|
|
|
|
Abstract Finding suitable cellulose degrading bacteria is the key to the effective application of cellulose in Inner Mongolia and other northern regions. In order to obtain some efficient cellulose-degrading bacteria, traditional isolation and enrichment culture technologies were used to isolate cellulose-degrading bacteria from the soil in western region of Inner Mongolia, and the degradation characteristics of cellulose-degradation bacteria were analyzed. A preliminary identification was based on analyzing the similarities of 16S rDNA gene sequences of the isolates. Cellulase activity was determined by DNS (3,5-dinitrosalicylic acid) method. And the enzyme-producing conditions of the highest enzyme activity cellulase-producing strain was further optimized. The results showed that a total of 59 cellulolytic bacteria were isolated, including 36 cold-adapted cellulose-degrading bacteria isolated at 10 ℃ and 23 cellulose-degrading bacteria isolated at 28 ℃. The 16S rDNA gene sequences indicated that these strains belonged to Alpha-Proteobacteria, Gamma-Proteobacteria, beta-Proteobacteria, Bacteroidetes, and Actinobacteria, respectively. Alpha-Proteobacteria was the first dominant group of room temperature cellulose-degrading bacteria, beta-Proteobacteria and Actinobacteria were the second dominant group. Gamma-Proteobacteria was the first dominant group of cold-adapted cellulose-degrading bacteria, Bacteroidetes was the second dominant group. At the genus level, Pseudochrobactrum was the first dominant genus of room temperature cellulose-degrading, while Pseudomonas was the second dominant genus, and Pseudomonas was the first dominant genus of cold-adapted cellulose degrading bacteria at low temperature, while Sphingobacterium was the second dominant genus. Among these strains, strain CB04 (belonging to Cellulomonas) had the highest carboxymethyl cellulose (CMC) activity. After optimization, the optimal conditions for enzyme production were as follows: the yeast extract was nitrogen source, the culture time was 4 d, the pH value was 7, and the CMC enzyme activity was up to 114.6 U/mL. The strain CB04 was a cellulase producing bacterium with great development potential. This study preliminarily revealed the composition and ability of cellulose-degradation bacteria in western region of Inner Mongolia, which provides a theoretical and practical basis for understanding and using cellulose-degradation bacteria for other applications.
|
|
Received: 14 April 2020
|
|
|
|
Corresponding Authors:
*chypapple@126.com
|
|
|
|
[1] 艾士奇, 赵一全, 孙志远, 等. 2018. 复合菌系降解纤维素过程中微生物群落结构的变化[J]. 生物工程学报, 34(11): 1794-1808.
(Ai S Q, Zhao Y Q, Zhao Z Y, et al.2018. Change of bacterial community structure during cellulose degradation by the microbial consortium[J]. Chinese Journal of Biotechnology, 34(11): 1794-1808.)
[2] 崔秀秀, 韩梅, 李丽娜, 等. 2016. 利用响应面法优化耐冷纤维素降解菌产内切纤维素酶的发酵条件[J]. 华中农业大学学报, 35(4): 62-69.
(Cui X X, Han M, Li L N, et al.2016. Optimizing fermentation conditions of endo-cellulase production by a psychrotrophic bacteria using response surface method[J]. Journal of Huazhong Agricultural University, 35(4): 62-69.)
[3] 戴芸芸, 钟卫鸿. 2016. 细菌降解木质纤维素的研究进展[J]. 化学与生物工程, 33(6): 11-16.
(Dai Y Y, Zhong W H.2016. Research progress on degradation of lignocellulose by bacteria[J]. Chemistry & Bioengineering, 33(6): 11-16.)
[4] 冯红梅, 秦永胜, 李筱帆, 等. 2016. 高温纤维素降解菌群筛选及产酶特性[J]. 环境科学, 37(4): 1546-1552.
(Feng H M, Qin Y S, Li X F.2016. Screening and enzyme production characteristics of thermophilic cellulase-producing strains[J]. Environmental Science , 37(4): 1546-1552.)
[5] 付丽, 朱红雨, 杜明楠, 等. 2018. 秸秆降解菌株的筛选、鉴定及生物学特性研究[J]. 中国农业大学学报, 23(12): 39-49.
(Fu L, Zhu H Y, Du M N, et al.2018. Screening, identification and biological characteristics of straw-degrading bacteria strains[J]. Journal of China Agricultural University, 23(12): 39-49.)
[6] 高星爱, 王鑫, 解娇, 等. 2020. 低温秸秆降解复合微生物菌剂的研究进展[J]. 生物技术通报, 36(4):144-150.
(Gao X A, Wang X, Xie J, et al.2020. Research progress on low temperature straw-degrading compound microbial agent[J]. Biotechnology Bulletin, 36(4): 144-150.)
[7] 姜立春, 赵丽萍, 林寿露, 等. 2017. 纤维素降解菌的筛选、鉴定与产酶条件优化试验[J]. 黑龙江畜牧兽医, 11: 166-172.
(Jiang L C, Zhao L P, Lin S L, et al.2017. Screening and identification of cellulose degrading bacteria and optimization of enzyme production conditions[J]. Heilongjiang Animal Science and Veterinary Medicine, 11: 166-172.)
[8] 李静, 张瀚能, 赵翀, 等. 2016. 高效纤维素降解菌分离筛选、复合菌系构建及秸秆降解效果分析[J]. 应用与环境生物学报, 22(4): 689-696.
(Li J, Zhang H N, Zhao C, et al.2016. Isolation and screening of cellulose decomposing microbe and the straw decomposing effect of complex microbial system[J] Chinese Journal of Applied and Environmental Biology, 22(4): 689-696.)
[9] 李淼淼, 沈飞, 张庆华. 2018. 纤维素降解复合菌系的微生物多样性及关键功能菌解析[J]. 环境科学学报, 38(7): 2690-2695.
(Li M M, Shen F, Zhang Q H.2018. Microbial diversity analysis of a cellulolytic microbial consortium and the key functional bacteria[J]. Acta Scientiae Circumstantiae, 38(7): 2690-2695.)
[10] 刘东阳, 王蒙蒙, 马磊, 等. 2014. 高效纤维素分解菌的分离筛选及其分解纤维素研究[J]. 南京农业大学学报, 37(6): 49-58.
(Liu D Y Wang M M, Ma L, et al.2014. Isolation of the efficient lignocelluloses degrading microbes and decomposition of cellulose[J]. Journal of Nanjing Agricultural University, 37(6): 49-58.)
[11] 罗立津, 万立, 陈宏, 等. 2015. 耐低温木质纤维素降解菌群的富集培养及其种群结构分析[J]. 农业生物技术学报, 23(6): 727-737.
(Luo L J, Wan L, Chen H, et al.2015. Enrichment culturing and bacterial community structures analysis of a cold-adapted lignocellulose degrading microflora[J]. Journal of Agricultural Biotechnology, 23(6): 727-737.)
[12] 马欣雨, 孙丽娜, 卢珊, 等. 2020. 秸秆降解菌的筛选及对秸秆的降解效果[J]. 生态学杂志, 39(4):1198-1205.
(Ma X Y, Sun L N, Lu S, et al.2020. Screening of straw degrading microbial strains and their degradation effects, 39(4): 1198-1205.)
[13] 潘虎, 董俊德, 卢向阳. 2012. 高效纤维素降解菌群的构建及其生物多样性分析[J]. 湖南农业大学学报(自然科学版), 38(2): 139-145.
(Pan H, Dong J D, Lu X Y.2012. Construction of a composite microbial system for efficient cellulose degradation and its biodiversity analysis[J]. Journal of Hunan Agricultural University(Natural Sciences), 38(2): 139-145.)
[14] 任继勤. 2018. 秸秆生物质能利用对节能减排的贡献潜力研究[J]. 北京交通大学学报(社会科学版), 17(4): 79-87.
(Ren J Q.2018. Simulation study on contribution potential of straw as biomass energy to energy conservation and emission reduction[J]. Journal of Beijing Jiaotong University (Social Sciences Edition), 17(4): 79-87.)
[15] 石祖梁, 刘璐璐, 王飞, 等. 2016. 我国农作物秸秆综合利用发展模式及政策建议[J]. 中国农业科技导报, 18(6): 16-22.
(Shi Z L, Liu L L, Wang F, et al.2016. Development model and policy proposal for comprehensive utilization of crop straw in China[J]. Journal of Agricultural Science and Technology, 18(6): 16-22.)
[16] 宋安东, 张百良, 吴坤, 等. 2005. 杂色云芝产木质纤维素酶及对稻草秸秆的降解[J]. 过程工程学报, 5(4): 414-419.
(Song A D, Zhang B L, Wu K, et al.2005. Production of lignocellulolytic enzymes and rice straw biodegradation by coriolus versicolor[J]. The Chinese Journal of Process Engineering, 5(4): 414-419.)
[17] 孙丽娜, 马欣雨, 刘克斌, 等. 2018. 秸秆的微生物处理处置及强化技术研究进展[J]. 沈阳大学学报(自然科学版), 30(3): 20-27.
(Sun L N, Ma X Y, Liu K B, et al.2018. A review on research advances on microbial treatment and strengthening techniques of crop straw[J]. Journal of Shenyang University(Natural Science), 30(3): 20-27.)
[18] 王海滨, 韩立荣, 冯俊涛, 等. 2015. 高效纤维素降解菌的筛选及复合菌系的构建[J]. 农业生物技术学报, 23(4): 421-431.
(Wang H B, Han Li- R, Feng J T, et al.2015. Screening of highly efficient cellulose degradation microbes and construction of composite strains[J]. Journal of Agricultural Biotechnology, 23(4): 421-431.)
[19] 王伟, 郑大浩, 杨超博, 等. 2019. 高效纤维素分解菌的分离及秸秆降解生物效应[J]. 中国农业科技导报, 21(8): 36-46.
(Wang W, Zhen D H, Yang C B, et al.2019. Isolation of high efficient cellulose decomposing bacteria and biological effects on straw degradation[J]. Journal of Agricultural Science and Technology, 21(8): 36-46.)
[20] 夏俊. 2018. 不同湿地土壤中真菌群落分析以及纤维素降解菌的筛选[J].生物化工, 4(4):37-39, 43.(Xia J. 2018. Analysis of fungal community structure and screening of cellulose degrading fungi in different wetland soils [J]. Biological Chemical Engineering, 4(4):37-39, 43.)
[21] 于慧娟, 郭夏丽. 2019. 秸秆降解菌的筛选及其纤维素降解性能的研究[J].生物技术通报, 35(2):58-63.
(Yu H J, Guo X L.2019. Screening of straw-degrading bacteria and study on their cellulose-degrading performances[J]. Biotechnology Bulletin, 35(2): 58-63.)
[22] 袁楠, 亢宗静, 卢圣鄂, 等. 2016. 富集培养下的若尔盖高原湿地低温纤维素降解细菌群落结构[J]. 22(3): 402-408 (Yuan N, Kang Z J, Lu S E, et al. 2016. Community structures of the cold-adapted cellulose-degrading bacteria in the Zoig(e) plateau wetland under enrichment culture conditions [J]. Chinese Journal of Applied & Environmental Biology, 22(3): 402-408.)
[23] 张梦君, 邱晨浩, 柴立伟, 等. 2019. 高效降解纤维素低温真菌的筛选、鉴定及发酵优化[J]. 微生物学通报, 46(10): 2494-2503.
(Zhang M J, Qiu C H, Cai L W, et al.2019. Screening, identification and fermentation optimization of cold-adapted fungi with high efficiency of cellulose degradation[J]. Microbiology China, 46(10): 2494-2503.)
[24] 张俙何, 洪春来, 朱凤香, 等. 2013. 农业废弃物资源化利用现状与前景展望[J]. 现代农业科技, 20: 209, 218.(Zhang X H, Hong C L, Zhu F X, et al. 2013. Current situation and prospect of agricultural waste resource utilization [J]. XianDai NongYe KeJi, 20: 209, 218.)
[25] 郑丽, 张海鹏, 宋艳培, 等. 2017. 纤维素降解菌的筛选、鉴定和糖化水平研究[J]. 广东农业科学, 44(2): 104-111.
(Zheng L, Zhang H P, Song Y P, et al.2017. Isolation, identification and saccharification level of cellulose degrading bacteria[J]. Guangdong Agricultural Sciences, 44(2): 104-111.)
[26] Behera B C, Parida S, Dutta S K, et al.2014. Isolation and identification of cellulose degrading bacteria from mangrove soil of Mahanadi River Delta and their cellulase production ability[J]. American Journal of Microbiological Research, 2(1): 41-46.
[27] Brown M E, Chang M C Y.2014. Exploring bacterial lignin degradation[J]. Current Opinion in Chemical Biology, 19: 1-7.
[28] Das A, Bhattacharya S, Murali L.2010. Production of cellulase from a thermophilic Bacillus sp. isolated from cow dung[J].American-Eurasian Journal of Agriculture Envi-ronment Science, 8(6): 685-691.
[29] Jönsson L J, Martín C.2015. Pretreatment of lignocellulose:formation of inhibitory by-products and strategies forminimizing their effects[J]. Bioresource Technology, 199: 103-112.
[30] Margesn R, Schnner F.1998. Low temperature bioremediation of a water contaminated with anionic surfactants and fuel oil[J]. Applied Microbiology Biotechnology, 49(4): 482-486.
[31] Ruijssenaars H J, Hartmans S.2004. A cloned Bacillus halodurans multicopper oxidase exhibiting alkaline laccase activity[J]. Applied Microbiology and Biotechnology, 65(2): 177-182.
[32] Selvam K, Senbagam D, Selvankumar T, et al.2017. Cellulase enzyme: Homology modeling, bindingsite identification and molecular docking[J]. Journal of Molecular Structure, 1150: 61-67.
[33] Wagg C, Schlaeppi K, Banerjee S, et al.2019. Fungal-bacterial diversity and microbiome complexity predict ecosystem functioning[J]. Nature Communication, 10: 4841.
[34] Wongwilaiwalin S, Laothanachareon T, Mhuantong W, et al.2013. Comparative metagenomic analysis of microcosm structures and lignocellulolytic enzyme systems of symbiotic biomass-degrading consortia[J]. Applied Microbiology and Biotechnology, 97(20): 8941-8954.
[35] Wu H, Dai X, Zhou S L, et al.2017. Ultrasound-assisted alkaline pretreatment for enhancing the enzymatic hydrolysis of rice straw by using the heat energy dissipated from ultrasonication[J]. Bioresource Technology, 241: 70-74.
[36] Xu W B, Huang T, Zhao J, et al.2015. Environmental conditions rather than microbial inoculum composition determine the bacterial composition, microbial biomass and enzymatic activity of reconstructed soil microbial communities[J]. Soil Biology and Biochemistry, 90: 10-18. |
|
|
|
